Recent studies have illuminated an important role for mitochondria in the expression of injury mediated by a number of distinct neurotoxic mechanisms. This suggests that the study of both the normal and pathophysiological function of mitochondria is critical in the investigation of neurodegenerative mechanisms. The long-term aim of this project is to identify the mechanisms by which mitochondria contribute to neuronal injury, and to use this understanding to design novel neuroprotective approaches. We will undertake four aims in this project that are designed to characterize a newly recognized phenomenon of spontaneous mitochondrial changes (SMC's) in cultures of neurons, astrocytes and HT22 cells. We believe that these changes provide a valuable window into mitochondrial function in normal neural cells. However, the mechanistic basis for SMC's is completely unknown. In aim 1 we will determine whether mitochondrial movement contributes to SMC's and develop novel imaging approaches to measure relevant phenomena. In specific aims 2 and 3, using SMD's as a marker we will investigate the mechanisms underlying SMC's using manipulations that alter both physiological and pathophysiological parameters. Finally, in specific aim 4 we will establish an organotypic culture model of the degeneration of nigral neurons in Parkinson's disease (PD) using approaches that allow us to identify dopaminergic neurons prior to performing imaging experiments. We will then assess mitochondrial function in dopaminergic neurons exposed to toxins that produce lesions specific to nigral dopaminergic neurons. This project will provide new insights into several critical aspects of mitochondrial function in neurons and astrocytes that will illuminate their role in normal cell physiology as well as in neurodegeneration and will direct future studies that develop novel approaches for intervention in a range of acute and chronic neurodegenerative diseases.